In the field of biotechnology, nanotechnology and electrical engineering are being utilized to develop new-type electronic devices such as chips, sensors and actuators.1-4 As a result of this work, a new scientific area called molecular electronics has emerged. Molecular electronics involves the combination of biotechnology and nanotechnology for expansion into information technology.
Traditionally, protein molecules have been shown to perform important roles in various biological systems. However, protein molecules can now also be used as an important element in human-designed electronic devices such as biosensors, transistors or memory devices. In addition, several hypotheses have been proposed for the design of electronic devices that are based on organic molecules and biomolecules.5-10 However, the devices developed in these studies only displayed a simple function and could not perform multiple functions.
Previously, the inventors of the present disclosure developed a simple electronic device that consisted of biomolecular hetero Langmuir-Blodgett films. This device displayed switching functions and could be used as a molecular diode.11,12 After the development of this initial device, the inventors of the present disclosure studied electroactive systems in regard to the kinetics of electron transport for further applications such as biosensors and bioelectronic devices. More recently, the inventors of the present disclosure focused on the development of various electrochemical-based biomemory devices that consist of cysteine-modified azurin and cytochrome c.17,18 However, the proposed biomemory device only displayed one function and only one bit information could be controlled. Therefore, the inventors of the present disclosure attempted to overcome the limitation of current bioelectronic devices, which do not have the ability to perform multiple functions. The biomemory chip of the present disclosure can perform multiple functions since it can exist in 2-state and 3-state (WRER-type and WORM-type).
In the present disclosure, a cysteine-modified azurin was designed using the site-directed mutagenesis technique and 4 different metal ions were introduced to make a 4-bit biomemory chip capable of performing multiple functions. The recombinant protein containing a cysteine residue and different metal substituents was designed and directly immobilized on gold surface without any chemical linkers. The redox property of the 4 different azurin variants (Co-substituted type, Ni-substituted type, Fe-substituted type and Mn-substituted type) was measured. The presence of the different metal substituents in the recombinant azurin protein was confirmed by UV-VIS spectroscopy. In addition, the surface morphology of the device was investigated by atomic force microscopy (AFM). The electrochemical properties of the 4 different azurin variants were assessed by cyclic voltammetry (CV) and open-circuit potential. In this analysis, the recombinant azurin variants were shown to be successfully modified by metal ion substitution. The natural property of each material and coordinated metal and electron transfer kinetics between the material and the substrate were the reason of the change in redox properties. As memory elements, the substituted metal ions could be used to store different information because they have different conducting states. In addition, the inflow and outflow of electrons could be easily controlled by applying an external potential. The memory functions of this device were verified by chronoamperometry (CA) and open-circuit potential amperometry (OCPA). In their previous study, the inventors of the present disclosure proposed the basic concept of this biomemory device.17 In the present disclosure, they confirmed the electrochemical properties of each modified azurin variant and developed a multifunctional 4-bit bio memory device using the recombinant azurin variants. If the characteristics of the different azurin variants were applied and integrated, a multifunctional biomolecule memory device would be realized. The control of immobilization of each azurin variant on the micro scale will be a vital factor. A computerized multi-channel electrochemical workstation is one possible application of the device of the present disclosure. It acquires various electrochemical data from each channel at the time and displays the results in parallel. This will be compared with a parallel result from each device and then integrated for operation as an independent working memory cell. FIG. 1 shows a schematic diagram of the 4-bit biomemory chip containing the recombinant azurin variants and its mechanism.
Throughout the specification, a number of publications and patent documents are referred to and cited. The disclosure of the cited publications and patent documents is incorporated herein by reference in its entirety to more clearly describe the state of the related art and the present disclosure.